Acute respiratory distress syndrome (ARDS) is a major cause of morbidity and mortality. Of the many potential predisposing factors, sepsis and pneumonia represent the two main causes of ARDS. In spite of an increase in survival in recent years mortality in patients with ARDS is still estimated around 30 to 40%. In this context, development of effective preventive strategies in patients at high risk of development of ARDS is of paramount importance. Unfortunately, the results of studies evaluating prophylactic regimens for ARDS have been mostly disappointing. The gaseous molecule carbon monoxide (CO) has been traditionally viewed as a toxic metabolic and industrial waste. However, recent studies have demonstrated an important physiologic role of CO in many biological systems. Specifically, strong anti-inflammatory, anti-oxidant and anti-thrombotic effects of CO gas administration and heme oxygenase activation (the enzyme that generates endogenous CO gas) have been demonstrated in several animal models. Previous studies conducted in our department have demonstrated that bronchoscopic instillation of endotoxin (LPS) in healthy volunteers elicits a compartmentalized pulmonary inflammatory response, serving as an excellent model to evaluate interventions directed towards suppression of lung inflammation at its earliest stages. In the current single blinded, randomized, placebo controlled study, we evaluated the effects of inhaled carbon monoxide on local pulmonary inflammatory responses following endotoxin administration. Twenty healthy subjects, male or female, age 18 to 40 underwent local endotoxin instillation, breathed CO or room air through a mask for 6 hours, and then a repeat bronchoscopy with lavage was done at 6 hours to assess the ability of CO to suppress local inflammation in the lung. We began enrolling subjects in May 2005 into the pilot study and completed the pilot study in October 2005. The pilot study enrolled 4 males and 4 females. We analyzed the data from the pilot and found the CO inhalation was well tolerated and did not result in any adverse events. In addition, at this low dose, no significant anti-inflammatory effects of CO inhalation were demonstrated. We then enrolled 12 males and 12 females in the main study. Our results from the main study demonstrate that endotoxin instillation and carbon monoxide inhalation were well tolerated and did not result in any adverse events. However, no significant anti-inflammatory effects of carbon monoxide on endotoxin induced inflammation were demonstrated. We have recently begun to explore the effects of CO inhalation on microRNA secretion by lung parenchymal cells using archived samples of bronchoalveolar lavage. The design of the current study allows us to assess the effects of short term CO exposure on normal lung airways and its interaction with local inflammation induced by endotoxin. Pilot studies show that microRNA species can be detected in bronchoalveolar lavage from normal and local airway inflammation. In addition, to further explore mechanisms, we are studying the differential expression of microRNA species in human bronchial epithelial cells grown in vitro in an air-liquid interface in order to assess mechanism associated with microRNA secretion. The project is now closed to enrollment and open for data and sample analysis only. During this year we transferred the samples and data to another open protocol where we will continue our data analysis and publication of the results.